Fig 1: Schematic representation of NEText, NT-3, and TrkC distribution in T cells of patients with schizophrenia and healthy controls. Legend: NT-3-mediated signal transduction pathways through dimerization of TrkC receptors, which results in an intracellular activation of extracellular signal-regulated kinases (ERKs), phosphatidylinositol-3-kinase (PI3K)-protein kinase B (AKT), and phospholipase C-? (PLC?) pathways promoting cell survival, proliferation, differentiation, synaptic plasticity, and gene expression in T cells. The molecular complex between the second extracellular loop of NEText (corresponding to amino acid residues 189–204) and NT-3 or TrkC seems to be essential for appropriate activation of the signaling cascade in a peripheric context that involves immune system cells. The red cross represents the mechanisms/pathways that are disrupted in schizophrenia.
Fig 2: Predictive results of the CABS-dock server. Legend: computational modelling of protein–peptide docking of NT-3 (1nt3:A) or TrkC (1wwc:A) and the peptide sequence (part of the second extracellular loop of NET: LLNGSVLGNHTKYSK). The docking prediction figures of proteins–peptide correspond to model 1. Clustering details of the ten models are shown in the respective tables.
Fig 3: Co-IPs of NEText and NT-3 or TrkC in T cells of patients with schizophrenia and healthy controls. Legend: Co-IPs of NEText, NT-3, and TrkC in T lymphocytes of patients with schizophrenia and controls. Legend: Measurement of NEText and TrkC levels in T cells and NT-3 levels in plasma, and their respective scatter plots and bar graphs (mean ± SD). (a) Co-IP NEText-blot NT-3 (12 patients with schizophrenia vs. 6 controls; p = 0.0003) and Co-IP NEText-blot TrkC (10 patients with schizophrenia vs. 6 controls p = 0.0020). (b) Co-IP NT-3-blot TrkC (12 patients with schizophrenia vs. 6 controls; p > 0.05) and Co-IP NT-3-blot NEText (12 patients with schizophrenia vs. 6 controls; p = 0.0021). (c) Co-IP TrkC-blot NT-3 (12 patients with schizophrenia vs. 6 controls; p > 0.05) and Co-IP TrkC-blot NEText (9 patients with schizophrenia vs. 6 controls; p = 0.030) (* p < 0.05; ** p < 0.01 and *** p < 0.001).
Fig 4: Levels of NET and TrkC in T cells, and NT-3 in plasma. Legend: quantification of NET and TrkC levels in T cells and NT-3 in plasma, and their respective scatter plots and bar graphs (mean ± SD). (a) NEText (45 patients with schizophrenia vs. 31 controls) and TrkC (31 patients with schizophrenia vs. 22 controls) levels were measured by Western blot and normalized with the housekeeping protein ß-actin. Relative units refer to the mean of control values after normalization. The results showed a reduction in NET levels (p < 0.0001) and TrkC levels (p = 0.0032) in T cells of patients with schizophrenia compared to the control group. (b) The analysis of NT-3 (54 patients with schizophrenia and 54 healthy controls) in plasma revealed a reduction in this neurotrophin in the plasma of patients in comparison with controls (p = 0.0040), whose concentration was measured by ELISA using a polynomial regression method (Y = absorbance; X = concentration, degree = 3; R2 = 0.9999).
Fig 5: Fluorescence microscopy images of blood T lymphocytes. Legend: fluorescence microscopy images of blood T lymphocytes. T cells were stained with anti-NEText + anti-Alexa Fluor 488 (green color) and anti-NT-3 + anti-Alexa Fluor 594 (red color) antibodies; or anti-NEText + anti-Alexa Fluor 488 (green color) and anti-TrkC + anti-Alexa Fluor 594 (red color) and expanded from peripheral blood mononuclear cells with an HCX PL APO CS 63.0 × 1.40 OIL UV objective. The white bar represents 10 µm. (a) NEText–NT-3 staining of T lymphocytes of patients with schizophrenia and controls. (b–d) Intensity profile graphs (distance in pixels vs. intensity in grey-scale values) of T cells of patients with schizophrenia (red) and controls (green): (c) distribution of NEText, (d) distribution of NT-3, and (e) distribution of merged results. NEText–TrkC staining of T lymphocytes of patients with schizophrenia and controls. (f–h) Intensity profile graphs (distance in pixels vs. intensity in grey-scale values) of T cells of patients with schizophrenia (red) and controls (green): (f) distribution of NEText, (g) distribution of TrkC, and (h) distribution of merged results.
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